Development of a ³¹P magnetic resonance spectroscopy technique to quantify NADH and NAD⁺ at 3 T

NADH and NAD⁺ act as electron donors and acceptors and NAD⁺ was shown to stimulate mitochondrial biogenesis and metabolic health. We here develop a non-invasive Phosphorous Magnetic Resonance Spectroscopy (³¹P-MRS) method to quantify these metabolites in human skeletal muscle on a clinical 3 T MRI scanner. This new MR-sequence enables NADH and NAD+ quantification by suppressing α-ATP signal, normally overlapping with NADH and NAD⁺. The sequence is based on a double spin echo in combination with a modified z-Filter achieving strong α-ATP suppression with little effect on NAD⁺ and NADH. Here we test and validate it in phantoms and in humans by measuring reproducibility and detecting a physiological decrease in NAD⁺ and increase in NADH induced by ischemia. Furthermore, the ³¹P-MRS outcomes are compared to analysis in biopsies. Additionally, we show higher NAD⁺ and lower NADH content in physically active older adults compared to sedentary individuals, reflecting increased metabolic health.